| 改进的质谱兼容的胶内酶切方法 |
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| 引用本文： | 王鸿丽,刘炳玉,谷苗,刘锋,吴胜明,张学敏,魏开华. 改进的质谱兼容的胶内酶切方法[J]. 军事医学科学院院刊, 2005, 29(2): 168-172 |
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| 作者姓名： | 王鸿丽 刘炳玉 谷苗 刘锋 吴胜明 张学敏 魏开华 |
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| 作者单位： | 军事医学科学院国家生物医学分析中心,北京,100850 |
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| 基金项目： | 国家“863”计划发展项目(2001AA233041) |
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| 摘 要： | 目的：改善胶内酶切的肽段回收率，提高蛋白质的有效鉴定率。方法：将改进的胶内酶切“四步提取法”，应用到96个双向电泳蛋白质斑点分析。结果：该方法提高了胶内蛋白质酶切肽段的回收率，改善了质谱图谱的质量。蛋白质的有效鉴定率达89．6％，其中，数据库检索得分80分以上者达82％。另外，证实了酶解原液直接用于质谱分析可获得较好信噪比的谱图。结论：应用“四步提取法”使得酶解肽段提取率和蛋白质的有效鉴定率明显提高。
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| 关 键 词： | 胶内酶切 四步提取法 肽质量指纹图谱 蛋白质组学 光谱分析，质量 内肽酶类 |
| 文章编号： | 1000-5501(2005)02-0168-05 |
Improved in-gel digestion for mass spectrometry |
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| WANG Hong-Li,LIU Bing-yu,GU Miao,LIU Feng,WU Sheng-Ming,ZHANG Xue-Min,WEI Kai-Hua. Improved in-gel digestion for mass spectrometry[J]. Bulletin of the Academy of Military Medical Sciences, 2005, 29(2): 168-172 |
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| Authors: | WANG Hong-Li LIU Bing-yu GU Miao LIU Feng WU Sheng-Ming ZHANG Xue-Min WEI Kai-Hua |
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| Affiliation: | WANG Hong-Li,LIU Bing-Yu,GU Miao,LIU Feng,WU Sheng-Ming,ZHANG Xue-Min,WEI Kai-Hua * |
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| Abstract: | Objective: To improve the peptide recovery during in-gel digestion. Methods and Results: Details of the "four-step extraction" of in-gel digestion were discussed in this paper. 96 of the protein spots from two dimensional gel electrophoresis were processed for evaluating this protocol. Results showed that this protocol enhanced the peptide recovery during in-gel digestion and improved the quality of MALDI-TOF-MS peptide mass fingerprinting (PMF). About 89.6% of protein spots were identified and 82% of the identified proteins obtained the scores of higher than 80 by MASCOT search engine. Also, direct PMF analysis of the digestion resulting solution was studied. Conclusion: The application of " four-step extraction" method enhances the peptide recovery during in-gel digestion,and increases identification of the protein. |
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| Keywords: | in-gel digestion four-step extraction peptide mass fingerprinting proteomics spectum analysis mass endopeptidases |
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